R Notebook

Data annotation - Merge clusters

library(Seurat)

Registered S3 method overwritten by 'data.table':
  method           from
  print.data.table     
Registered S3 method overwritten by 'htmlwidgets':
  method           from         
  print.htmlwidget tools:rstudio
Seurat v4 was just loaded with SeuratObject v5; disabling v5 assays and validation
routines, and ensuring assays work in strict v3/v4 compatibility mode

library(tidyverse)

── Attaching core tidyverse packages ────────────────────────────────────────── tidyverse 2.0.0 ──
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ℹ Use the conflicted package (<http://conflicted.r-lib.org/>) to force all conflicts to become errors

library(CelltypeR)

Warning: replacing previous import ‘data.table::last’ by ‘dplyr::last’ when loading ‘CelltypeR’
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Attaching package: ‘CelltypeR’

The following object is masked from ‘package:ggplot2’:

    annotate

Read in the step7 object

nsc <- readRDS("/Users/rhalenathomas/Documents/Data/scRNAseq/ADHD_ZNZ_Mcgill/ADHDresultsFeb10/NSC/step7/objs7/seu_step7.rds")

Warning message:
R graphics engine version 15 is not supported by this version of RStudio. The Plots tab will be disabled until a newer version of RStudio is installed. 

See the Dimplot

DimPlot(nsc, group.by = 'integrated_snn_res.0.25', label = TRUE) 

DimPlot(nsc, group.by = "Celltypes1", label = TRUE)

#DimPlot(nsc, group.by = "Celltypes2", label = TRUE)
#DimPlot(nsc, group.by = "CelltypesMain", label = TRUE)
DimPlot(nsc, group.by = "CelltypesMain2", label = TRUE)

#DimPlot(nsc, reduction = "umap", label = TRUE, pt.size = 0.1, raster = FALSE) 

NPC_glia <- FindMarkers(nsc, ident.1 = "NPC_glia", ident.2 = c("Progenitors", "NPC"), only.pos = TRUE)

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View(Neurons_sub)
View(NPC_markers)

Name Fibroblast as Glia Keep NPC-glia Change Precursor vs NPC

NSC-vas is now NSC-APOA1 NSC- SOX2

NPC-MEF2C Progen is really NPC-POU5F1 NPC-S100B

Neurons-DCX (was neurons) Neurons-GRIA1 (was excitatory neurons)

See the neurons object

fcn <- readRDS("/Users/rhalenathomas/Documents/Data/scRNAseq/ADHD_ZNZ_Mcgill/ADHDresultsFeb10/FCN/step7/objs7/seu_step7.rds")

Transfer lables predictions

need to match version 4 and version 5 seurat

Myanchors <- FindTransferAnchors(reference = MBO ,query = fcn, dims = 1:30)

Performing PCA on the provided reference using 1076 features as input.
Projecting cell embeddings
Error in UseMethod(generic = "GetAssayData", object = object) : 
  no applicable method for 'GetAssayData' applied to an object of class "Assay5"

Visualize expression

proliferation = c("PCNA","MKI67")
neuralstem = c("SOX2","NES","PAX6","MASH1")

DotPlot(fcn, features = proliferation, group.by = "integrated_snn_res.0.25")

FeaturePlot(fcn, features = proliferation)

neuralstem = c("SOX2","NES","PAX6","MASH1")

DotPlot(fcn, features = neuralstem, group.by = "integrated_snn_res.0.25")

Warning in FetchData.Seurat(object = object, vars = features, cells = cells) :
  The following requested variables were not found: MASH1

FeaturePlot(fcn, features = neuralstem)

Warning in FetchData.Seurat(object = object, vars = c(dims, "ident", features),  :
  The following requested variables were not found: MASH1

# simplify cell types 1
cluster.ids<-c("Differentating_to_neurons", "NSC", "NSC", "Neurons", "Endothelial", "Endothelial", "Endothelial", "NSC", "Neuroblasts", "Neurons") 
  Idents(fcn) <- 'integrated_snn_res.0.25'
  names(cluster.ids) <- levels(fcn)
  fcn <- RenameIdents(fcn, cluster.ids)
  fcn <- AddMetaData(object=fcn, metadata=Idents(fcn), col.name = "CelltypesMain1")

DimPlot(fcn, label = TRUE, group.by = "CelltypesMain1")

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